UA34441C2 - Pharmaceutical preparation for reducing concentration of glucose in blood - Google Patents

Abstract

The invention relates to a method of reducing blood glucose levels in humans, particularly those afflicted with diabetes.

Description

The invention relates to a method of reducing glucose content! in the blood of people, especially those with diabetes.

Diabetes mellitus is a systemic disease characterized by a violation of the action of insulin and other regulatory hormones in the metabolism of carbohydrates, fats and proteins, as well as in the structure and function of blood vessels. The primary symptom of diabetes is hyperglycemia, often accompanied by glucosuria (presence of a large amount of glucose in the urine) and polyuria (excretion of a large volume of urine). An additional symptom! appear in chronic and long-term diabetes. This is a symptom! include degeneration of the walls of blood vessels. Although many different organs are affected by such changes in blood vessels, as it turned out, nerves, eyes and kidneys are the most susceptible! As such, long-term diabetes, even when treated with insulin, leads to blindness.

Lime! two types of diabetes. Diabetes; | type are recognized as youth, connect! with ketosis, develop early in life, have severe symptoms and, in the near future, vascular damage. Regulation of this type of diabetes is difficult and requires exogenous insulin administration. Diabetes mellitus! type II characteristic of adults, stable! to ketosis, develop in the late period of life, proceed more gently and more slowly.

One of the most significant successes in the history of medical science came in 1922, when

Bznting and Best demonstrated therapeutic effects! insulin in diabetic dogs.

However, even today, a clear picture of the main biochemical disorders caused by the disease, gum disease and diabetes remain an important problem of medicine! It is estimated that 295% of the US population is affected by various forms of diabetes. Oral administration of hypoglycemic agents has become an important achievement in the treatment of hyperglycemia. Oral hyperglycemic agents are commonly used to treat diabetes in adults. Observation of glucose metabolism in model animals and in people with type II diabetes suggests that sexual steroids play a decisive role in the phenotypic expression of hyperglycemia. These observations prompted the study of the influence of androgens and estrogens on the level of glucose! in the blood Administration of testosterone to normal female rats or ovariectomized female rats leads to marked resistance to insulin, which correlates with morphological changes in muscles, Holman et al., At. in. RPpuvio!., 259, E555-560 (1990); ibid., 262, E851-855 (1992). In streptozotocin diabetic rats, implanted testosterone antagonizes the ability of residual insulin to maintain glycemic control, Le et al., Epioshypoiodu, 116, 2450-2455 (1985).

On the contrary, glucosuria disappears in castrated diabetic CC muscles and reappears when androgens are substituted in these muscles, Nonaka et al. Urp. in). May 5si., 50, 1121-1123 (1988); Hyguychy et al., Ehr.

Apit., 38, 25-29 (1989).

The result! administration of estrogens also supports the hypothesis that the balance between androgens and estrogens is a decisive factor for the development of hyperglycemia. Daily administration of estradiol to diabetic CK muscles normalizes glucose levels! in the blood and eliminates glycosuria, Toshiro et al., Urp. .). May 5si., 51, 823-826 (1989). Estradiol also lowers the level of glucose in the blood in mice C57ВИ /62-0Бб/0р, Dubuk, Rgos. boss. Ex. Axes Med., 180, 468-473 (1985) and S57VI /Kv.-yr/d0, Harris,

Apayuptis! VNesoga, 225, 310-317 (1989).

The present invention offers a way to reduce the concentration of glucose in the blood of mammals that need a reduced concentration of glucose! in the blood, which consists in the introduction of a therapeutically effective amount of an anti-estrogen compound or its pharmaceutically acceptable salt or solvate.

The term "anti-estrogen", used in this description and claims, means a non-endogenic, non-steroidal chemical compound that exhibits anti-estrogenic activity (as further defined below), but at the same time possesses hormonal or estrogenic activity in some selective tissues and organs. Historically, the literature speaks of anti-estrogens as compounds that bind to estrogen receptors (for example, compete with estradiol for these receptors) and inhibit the response of the uterus or breast to estrogen, etc. Many of these compounds, as it was shown, apply! as means against fertilization, anticancer means or means against other pathological conditions for which inhibition of estrogen activity is useful. Many of these compounds, as noted, are agonist-antagonist mixtures, that is, at a certain dose level, they behave as strogen agonists, while at higher doses they are strogen antagonists. The ability to determine classic anti-estrogens is good! to general specialists, for example, descriptions in the work: Khzyis et al., U.A. Epdoshypoiodu, 1981, 108, 164-172.

Recently, it was discovered that some chemical classes of historically established anti-estrogens have the ability to act favorably on some tissues and organs that react to estrogen, while being recognized as antagonists of the estrogen reaction in others (tissues and organs). It is not clear whether this difference can be explained by the quantitative effect of the agonist-antagonist mixture or by a qualitatively different mechanism of action. An example of a favorable differential effect is Raloxifene, which has the ability to reduce the level of serum lipids and reduce bone resorption without unwanted uterine effects in postmenopausal women.

It should be noted that not all compounds that can act as anti-estrogens according to the classical definition made above can have the activity and utility described in the invention.

In general, the anti-inflammatory compounds claimed in this invention are those compounds which are anti-inflammatory in the classical sense and which have useful properties

genetic properties. The most common compounds are the chemical class of triarylethylene or 7-triarylpropenonamine!. Other anti-aging chemical class! are based on their merits.

The preferred and most preferred compounds of this invention are those which demonstrate the greatest hypoglycemic effect and which also show the least undesirable strangulating side effect in organs such as the breast and uterus, Jones et al., U. Med.

Spent 1984, 27, 1057-1066. In addition, preferred compounds of the invention, which have very low activity of strogen agonists, must! It is most useful when treating male individuals to avoid side effects of feminization, such as gynecomastia.

Antistrogens, considered within the framework of the present invention, are compounds that have an obvious binding affinity to strogen receptors, generally denoted by the value

Ki, the value of which is greater than 0.05 nM and less than 5000 nM, calculated from the inhibitory values! ISvo with the help of the equation Ki-xISvo/-((/kKa)|, where Г is the concentration of the labeled ligand and Ka is the dissociation constant of the ligand-receptor complex, determined under the conditions of saturation or from inhibition of binding of the "cold" ligand.

Methods of performing a binding analysis to determine the anti-strogenic and anti-strogenic activity of lime! to specialists, for example, Blok and Hood, Mebsiepsev5, 26, 1453-1458 (1980);

Block and Hood, Epidemiology, 109, 987-989 (1981); Blok et al., Mebsieepsev, 32, 1031-1036 (1983). Also known is the relationship between the apparent (relative) binding affinity and the inhibitory value of ISvo, the concentration of the labeled ligand, and the dissociation of the ligand-receptor complex, given in the above formula.

The preferred method for determining estrogenic/antiestrogenic activity is described in the methods below.

In the present invention of sentences! next class! anti-estrogens, which are believed to be useful in the methods of the invention: 1) triarylethylene; 2) 2,3-diaryl-2H-1-benzopyran; 3) 1-aminoalkyl-2-phenylindole; 4) 2-phenyl-3-aroylbenzthiophene!; 5) 1-substituted-2-aryldihydronaphthalene!; and 6) 2-substituted-3-arylbenzofuran!. Each of these classes is described in detail below.

The general chemical term used in the description of the compounds of this invention has its ordinary meaning. For example, the term "alkyl" by itself or as part of a second substituent denotes an alkyl radical with a normal or branched chain having a fixed number of carbon atoms, such as methyl, ethyl, propyl, isopropyl, and higher homologues and isomers, if indicated.

The term "Alkoxy" means an alkyl with a fixed number of carbon atoms, connected to the main molecule by an oxygen atom, for example, a methoxy-, ztoxy-, propoxy-, butoxy-, pentyloxy- and hexyloxy group, and also includes the structure! with a branched chain, for example, isopropoxy and isobutoxy groups.

The term "Ci-C7-alkanoyloxy" denotes the group -0-C(0)-BH2, where Ka is hydrogen or Ci-Cv-alkyl and includes formyloxy-, acetoxy-, propanoyloxy-, butanoyloxy-, pentanoyloxy-, and hexanoyloxy groups and they like groups! and also includes the isomer with a branched chain, for example, 2,2-dimethylpropanoyloxy- and 3,3-dimethylbutanoyloxy group.

Similarly, the term "C--C7-cycloalkanoyloxy" denotes the group -O-C(0)-C3-Cv-cycloalkyl, where

C3-C6-cycloalkyl includes cyclopropyl, cyclobutyl, cyclopentyl and cyclohexyl.

The term "Ci-C-Alkoxy-Ci-C7-alkanoyloxy" denotes the group -O-C(O)-H85-O-C1-Ci-alkyl, where P? - bond, Ci-Cv-aluoxycarbonyloxy group or Ci-Cv-alkanediyl, and includes, for example, methoxycarbonyloxy-, ztoxiccarbonyloxy-, propoxycarbonyloxy-, butoxycarbonyloxy-, methoxyacetoxy-, methoxypropanoyloxy-, methoxybutanoyloxy-, methoxypentanoyloxy-, methoxyhexanoyloxy- , ztoxyacetoxy-, ztoxypropanoyloxy-, ztoxybutanoyloxy-, ztoxypentanoyloxy-, ztoxyhexanoyloxy-, propoxyacetoxy-, propoxypropanoyloxy- and propoxybutanoyloxy groups and similar groups!.

The term "unsubstituted or substituted aroyloxy" denotes the group -O-C(O)-aryl, where aryl is phenyl, naphthyl, thienyl or furyl, and each group is unsubstituted or monosubstituted by a hydroxy group, halogen, C1-C3-alkyl or Ci-C3-Alkoxy group. x The term "halogen" means chlorine, fluorine, bromine or iodine. x The term "unsubstituted or substituted arylcosycarbonyloxy" denotes the group -0- -C(0)-O-aryl, where aryl is a phenyl, naphthyl, thienyl or furyl group, and each group is substituted or unsubstituted by a hydroxy group, halogen, C1-C3-alkyl or Ci-C3-Alkoxy.

The term "pharmaceutically acceptable salts" refers to salts of compounds of the above classes, which are practically non-toxic! for living organisms. Typical pharmaceutically acceptable salts include salts obtained by reacting compounds of the above classes with pharmaceutically acceptable inorganic or organic acids or pharmaceutically acceptable alkali metals or organic bases, depending on the type of substituent present in the compound.

Try it! pharmaceutically acceptable inorganic acids, which can be used to obtain pharmaceutically acceptable salts, include hydrochloric, phosphoric, sulfuric, hydrobromic, hydroiodic and phosphoric acids and similar acids. Try it! pharmaceutically acceptable organic acids that can be used to obtain pharmaceutically acceptable salts include aliphatic mono- and dicarboxylic acids, oxalic, carbonic, citric and succinic acids, phenyl-substituted alkanoic acids, aliphatic and aromatic sulfonic acids and other acids. Pharmaceutically acceptable salts obtained from inorganic and organic acids include, for example, hydrochloride, bromide, nitrate, sulfate,

pyrosulfate, bisulfate, sulfite, bisulfite, phosphate, monohydrophosphate, dihydrophosphate, metaphosphate, pyrophosphate, iodohydrate, fluorohydrate, acetate, propionate, formate, oxalate, citrate, lactate, p-toluenesulfonate, methanesulfonate, maleate and others

Many compounds from the above classes, which contain a carboxy group, a carbonyl group, a hydroxy group, and a sulfoxide group, can be converted into pharmaceutically acceptable salts by reaction with a pharmaceutically acceptable alkali metal or organic base. Try it! pharmaceutically acceptable organic bases that can be used to obtain pharmaceutically acceptable salts include ammonia, amines such as triztanolamine, triztylamine, ztylamine and others. Examples of pharmaceutically acceptable base alkali metals include compounds of formulas! MO, where M is an alkali metal, for example, sodium, potassium or cast iron, and 2 is hydrogen or C-C4-alkyl.

It should be understood that an individually taken anion or cation forming part of any salt of the invention is not dangerous, as long as the salt as a whole is pharmacologically acceptable and the anion or cation does not introduce an undesirable quality.

In addition, some of the compounds proposed as useful in the methods of the present invention can form solvates with water or common organic solvents. Such solvates are within the scope of the present invention.

The first class of anti-estrogens is triaryl ethylene. These compounds are widely known, proposed and obtained by methods described in US patents 4,536,516 and 2,914,563; Ogawa and others,

Spent React. Vii., 39 (4), 911 (1991), which includes everything! as villages. Typical representative compounds of this class include Tamoxifen, Clomiphene and (2)-4-11-(4--2-dimethylamino)ztoxy)phenyl-2-(4-isopropylphenyl)-1-butenylphenyl monophosphate.

Triarylethylenes include compounds of formula I

IN

-- and

KO) eens

In u s, v i in which EK is the main zfir group of formulas! -OSyaNapA, where n: 2, C or 4 and A is a dialkylamino group, where alkyl contains 1-4 carbon atoms, or a cyclic group selected from M-piperidinyl, M-pyrrolidinyl, M-morpholinyl and M-hexamethyleneimino groups!; each of B! is independently hydrogen, a hydroxy group, a halogen or a methoxy group, and their pharmaceutically acceptable salts and solvates!.

US patent Me 4536516 describes Tamoxifen - triarylethylene of the formula Ia (n3zs) amscany c-syusnz and its pharmaceutically acceptable salts of addition of acids and solvates and the method of its preparation.

Also, US patent No. 2,914,563 describes triaryl ethylenic formulas. HER, )54

Xx 1

Y Uai s-s / rya i, and ro

Kk II in which K is the main zfir group of formulas! -OSoNapA, where n is 2, C or 4 and A is a dialkylamino group, where the alkyl groups independently contain 1-4 carbon atoms, or a cyclic group such as M-piperidinyl, M-pyrrolidinyl or M-morpholinyl. The -OSyaNopA group is bonded to phenyl in the n-position relative to the carbon bonded to xylene. Each of B! independently appear as hydrogen, hydroxygroup

poi, halogen or methoxy group; X is a halogen, and their pharmaceutically acceptable salts and solvates. There are also suggestions! obtaining such compounds.

Ogawa et al. proposed the triaryl ethylenic formula ПІ, о-сносно-МвгВ3 and снсН3 in ІіІІ in which В: and ВЗ are independently selected from hydrogen or methyl; 7 - isopropyl, isopropen-2-yl, mono- or dihydroxyisopropyl; VZ - hydroxy group or phosphate (-ОРОзНг»г), and their pharmaceutically acceptable salts and solvates!. This article also suggests a method of obtaining such compounds.

The second class of anti-estrogens is 2,3-diaryl-2H-1-benzopyran. The connection of proposals and the methods described in EP 470 Z10A1 and in the articles of Sharma et al., U. Med. Spent 33, 3210, 3216, 3222 (1990), which included! as a link in its entirety. Typical compounds of this class include 2-(4-(2-(1-piperidinyl)zoxy|phenyl)-3-(4-hydroxyphenyl)-2H-1-benzopyran; 2-(4- /2-(1- piperidinyl)zthoxy/phenyl)-3-phenyl-7-methoxy-2IN-1-benzopyran; 2-(4-/2-(1-piperidinyl)zthoxy/phenyl)-3-(4-hydroxyphenyl)-7- hydroxy-2H-1-benzopyran.

EP 470310A1 describes the benzopyrany formulas of IM,

F. Y

Y and F

In the one in which R and B" (can be the same or different) - hydrogen, a hydroxy group, a C1-C7-alkoxy group or a C2-C8-alkoxycarbonyl; solvate, as well as the method of obtaining them.

The third class of anti-estrogens is 1-aminoalkyl-2-phenylindol. Join offers and receipts! according to methods, descriptions by von Angerer and others, y). Honey. Spet., 33, 2635 (1990), which is incorporated herein by reference in its entirety. 1-Aminoalkyl-2-phenylindole, described by von Angerer et al., i.e. compounds of the formula M, in

Where is the KE? - hydrogen or methyl; K!9 and V!!! - methoxy- or hydroxy group; Monday to Friday 4 to 8; M - group

MA2B"Z, where "2 and B!Z are independently hydrogen, methyl or zyl or one of B? or BZ is hydrogen, and the second is benzyl, or together with a nitrogen atom they form pyrrolidinyl, piperidinyl, or morpholinyl, and their pharmaceutically acceptable salts and solvates!. Ability! synthesis of these lime compounds from links or suggestions! here

The fourth class of anti-estrogens is 2-phenyl-3-aroylbenzo/b/thiophene!; (2-triarylpropenonies). These compounds are proposed and obtained according to the methods described in US patents Nos. 4,133,814 and 4,418,068 and in the article by Jones et al., U. Med. Spet., 27, 1057-1066 (1984), which included! here as a link in its entirety. Typical compounds of this class include Raloxifene (b-hydroxy-2-(4-hydroxyphenyl)benzo/p/thien-3-yl4- (2-(1-piperidinyl)zthoxy|phenyl)methanone hydrochloride, formerly Keoxifen and/b -hydroxy-2-(4-hydroxyphenyl)-benzo/p/thien-3-yl/-/4-(2-/1-pyrrolidinyl/zthoxy)phenyl/methanone hydrochloride. 2-Phenyl-3-aroylbenzo/b/ Thiophenes are shown in US patent No. 4133814 and have the formula MI, (o; p-o-

FU Sun.

V!b 5

IN!

U1 in which ВЭ is hydrogen, hydroxy-, C-C7-alkoxy, C-C7-alkanoyloxy-, C3-C7-cycloalkanoyloxy-, C-C7-alkoxy- C-C7-alkanoyloxy-, substituted or unsubstituted aroyloxy- or a substituted or unsubstituted aryloxycarbonyloxy group;

B!" - hydrogen, chlorine, bromine, hydroxy-, C-C-Alkoxy-, adamanthoyloxy-, C-C7-alkanoyloxy-, C3-C7-cycloalkanoyloxy-, C-C-Alkoxy-Ci-C7-alkanoyloxy-, substituted or unsubstituted aroyloxy- or substituted or unsubstituted aryloxycarbonyloxy group;

B!8 - group -"ОСНаСНо-Х'-МА"Б2о, where X is a bond or -СНе-, BZ and 29 - b-С.4-alkyl or together with a nitrogen atom form pyrrolidinyl, piperidinyl, hexamethyleneiminyl or morpholinyl, and their pharmaceutically acceptable salts and solvates!.

Methods of obtaining these compounds are described in US patent No. 4133814. Raloxifene and the method of obtaining it are described in US patent No. 4418068.

The fifth class of anti-estrogens is 1-substituted-2-aryldihydronaphthalene!. These compounds were proposed and obtained according to the methods described in US patents Nos. 4400543, 4323707, 4230862 and 3274213, which are fully included! as a link. Typical compounds of this class are Nafoxyden and Trioxyfen.

An example of 1-substituted-2-aryldihydronaphthalene compounds! in US patent No. 4,230,862, which describes compounds of formula I, (9)

ІІ х

Ff and Ge; VIV 17 her MII in which 7 is СНо-СНе- or -СНУСН-; ВЭ - hydrogen, hydroxy- or Ci-Cv-alkoxy group; B!" is a hydrogen, chlorine, bromine, hydroxy-, C-C5-alkyloxy-, C-C5-acyloxy-, C-C5-alkoxycarbonyloxy-, benzyloxy- or adamanthoyloxy group; B! - a C-C5-alkyloxy group or group -"ОСН2о-СНе МВ'УВ2О, where "З and о are independently C-C4-alkyl or, together with the nitrogen atom with which they are connected, form pyrrolidinyl, piperidinyl, hexamethyleneimino group or morpholinyl, with with the restriction that if B" is hydrogen, then K!5 is hydrogen, a hydroxy- or a C-C5-alkyl group and at least one of B"? and B" differs from hydrogen, and their pharmaceutically acceptable acid addition salts and solvates, as well as the method of obtaining such compounds.

An example of 1-substituted-2-aryldihydronaphthalenes is also given in US Pat.

Mo 3274213, which describes the combination of formulas! MIP,

M: -o-SaNozd-MViZ820

of beer, and MITI in which BZ and 29 - C.-Cv-alkyl or together with the nitrogen atom with which they are connected, form a 5-7-membered saturated heterocyclic radical, selected from pyrrolidinyl, 2-methylpyrrolidinyl, 2, 2-dimethylpyrrolidinyl, piperazinyl, 4-methylpiperazinyl, 2,4-dimethylpiperazinyl, morpholinyl, piperidinyl, 2-methylpiperidinyl, 3-methylpiperidinyl, hexamethyleneiminyl, homopiperazinyl, and homomorpholinyl; d - 2-6; r - 1-4; IN! - C-C-Alkoxy group, and their pharmaceutically acceptable salts and solvates!. Ability! obtaining these compounds is described in the same patent.

The sixth class of anti-estrogens is a 2-substituted-3-arylbenzofuran. This is a combination of propositions and results based on the methods described in the article by Theo et al.). May Spet., 35, 1330-1339 (1992), which is included here as a reference. Proposals by Theo et al.,.). Honey. Spet.. Z5, 1330-1339 (1992) 2-substituted-3-arylbenzofurans include compounds of formulas IX, 22 ko o o r)-e osnosnov?! IX where X? - halogen; IN? - connection or methylene; 222 - hydrogen or methyl; 223 - group -МВ'ЗВ2?, where В" and 229 are independently Ci-Ca4-alkyl or, together with the nitrogen atom with which they are bonded, form pyrrolidinyl, piperidinyl, hexamethyleneiminyl or morpholinyl, and their pharmaceutically acceptable salts and solve!.

There are suggestions! way! obtaining those compounds.

A preferred class of compounds useful in the methods of the present invention are benzthiophene. The most preferred are benzothiophenes of the formula X, osnosna-xI--y! "C with B! days I 4 in which X! - connection or methylene; В!5 - hydroxy-, methoxy-, C-C7-alkanoyloxy-, C3-C7-cycloalkanoyloxy-, C1-C8-alkoxy-Ci-C7-alkanoyloxy-, substituted or unsubstituted aroyloxy-, or substituted or unsubstituted aryloxycarbonyloxy group; B!" - hydrogen, chlorine, bromine, hydroxy-, methoxy-, C-C7-alkanoyloxy-, C3-C7-cycloalkanoyloxy-, C1i-C8-alkoxy-Ci-C7-alkanoyloxy-, substituted or unsubstituted aroyloxy-, or a substituted or unsubstituted aryloxycarbonyloxy group; M! is a heterocyclic ring selected from pyrrolidinyl, piperidinyl, or hexamethyleneiminyl, and their pharmaceutically acceptable salts and solvates. Raloxifene and its pyrrolidinyl analog are especially preferred.

This invention offers a way to lower glucose levels! in the blood of mammals, resulting in the introduction of a therapeutically effective amount of an anti-estrogen compound or its pharmaceutically acceptable salt or solvate. The term "therapeutically effective amount", as used herein, means the amount of the compound necessary to achieve a hypoglycemic effect after administration, preferably to people suffering from or susceptible to diabetes in adults. The hypoglycemic effect considered in the present method includes both medical therapeutic and (or) prophylactic treatment. Also within the framework of the present invention, concomitant treatment with oral hypoglycemic agents, insulin or insulin derivatives is considered. Such additional therapeutic means of duty! to be determined by the attending physician taking into account the circumstances.

The hypoglycemic activity of the compounds of the present invention was determined by testing the effectiveness of ip mimo on viable male obese diabetic mice. The test methodology is described below.

We used 5-6-month-old male, viable yellow mice (MULLI-AU"a, with congenital diabetes mellitus, hereinafter referred to as diabetic mice) from the colonies of Lilly. The UU strain, carrying the AU"U mutation, was donated by Dr. Wolf to the research Lilla's laboratory about 20 years ago. The colony was supported by a sister x brother cross between the A" and a/a masses. Males!

their yellow muscles are obese, hyperglycemic, hyperinsulinemic and resistant to insulin.

Mice were housed 6 per plastic cage with litter, water, and ad libitum Purina Formulab 5008 (Purina Mills, St. Louis, MO). The temperature in the animal rooms was maintained at 23 - 220. The light in the animal rooms was 600--1800 hours.

Antistrogen! consumed in various doses as food additives. Each dose of antistrogen was tested on 6 mice contained in one cell. The compounds were mixed into the ground feed, which was then granulated. Control mice were given granulated food without any addictive compound. Blood samples were taken from the tail vein before the test itself and one week after the test started. Glucose concentration! glucose oxidase was determined in the blood! on a fast-flow analyzer 300

Alpkem (Klakzmaus, OK).

The numbers shown in tables 1-3 are the average values obtained in each round, while each table describes a separate round. Doz! are average doses based on actual food consumption and body weight. Table 2 describes the result!

ISI 164384 is an anti-estrogen that does not have estrogenic activity and falls outside the scope of the present invention. The statistical analysis of these whites is performed by the method of significant difference, based on the analysis of variations.

Table 1

Hypoglycemic activity of compounds tested on the muscles of obese people with diabetes

THREE ele Godney o | tdney | nicer

Tamoxifen 0.08 4834-15 280-442 288428 0.20 46631 1941429 196423 0.74 533316 24032 188520 2.30 450-339 183522 207426

Trioxyphene 0.09 " 483-420 223-426 234-332 0.27 5OT13 192427 164414 o 471432 186427 1584112 " 501-2 154-4 141-4

Control untreated meat 2 0.0 54331 457427 4349

Table 2

Hypoglycemic activity of compounds tested on the muscles of obese people with diabetes

THREE bzhlene God days | days | better/

CPI 164384 0.03 366-17 326421 317-300

M-p-butyl-M-methyl-11-(3,17 p-dihydroxystra-1,3,4(10)-trien-7o-yl)-undecamide 0.30 381-117 388-16 327-14 2 ,80 383527 292-268 297-415

IV-hydroxy-2-(4- 0.03 35413 161414 318.15 hydroxyphenyl)-benz(b|thien-3- yl(4-(2-1-pyrrolidinyl)zthoxy)| 0.24 phenyl|methanone hydrochloride 4 "5 327- 420 250-333 210-426

And 379-18 185444 163-332 0.02 17; - ztynylostradiol 400--17 108-533 10552

Table C

Hypoglycemic activity of compounds tested on the muscles of obese diabetics ooieltyenoya 07 odee | 00 Konientratsiitlyukuzne news

1 guilty 000 fugitive | alone | 00tlney | 00 ladneys

Raloxifene 0.01 36323 354--16 357-111 0.28 406420 378-417 31411 0.84 37334 239-439 196-935 0.24 407-414 23134 165-432 17 -estinyl estradiol 3 7.361 091-101 -426 17725 0.025 399410 186410 151414

The following methods describe the preferred methods for determining estrogenic/antiestrogenic activity of the compounds described above.

Femur density.

Seventy-five-day-old female Sprogue Dawley crosses (weight range: 225-275 g) were treated with

Charles River Laboratories (Portage, MI). Chris was kept in groups of Z and given as much food as he wanted (calcium content approximately 195) and water. The temperature in the room was maintained at 22.2 5 1.72C with a minimum relative humidity of 4095. The light period in the room was 12 hours of light and 12 hours of darkness.

One week after arrival, rats were subjected to bilateral ovariectomy (ovariectomy) under anesthesia (44 mg/kg Ketamine and 5 mg/kg Xylazine (Butler, Indianapolis, MI) intramuscularly).

Treatment with a carrier, estrogen, or test compound begins on the day of surgery after recovery from anesthesia. Oral dosing is carried out by tube feeding in 0.5 ml of 1956 carboxymethyl cellulose or 2095 cyclodextrin. The body mass is determined during the surgical operation and a few weeks later, and the dosage is adjusted to the change in mass! bodies Varicectomized (ovex) rats and non-varicectomized (intact) rats treated with vehicle or estrogen are evaluated in parallel in each group for negative and positive control.

Rats are treated daily for 35 days (6 rats per treated group) and killed by carbon dioxide asphyxiation on the 36th day. A period of time of 35 days is sufficient to maximally decrease bone density in ovariectomized rats, measured as described here. When dead, the uterus is removed, free from extraneous tissues, and the liquid content is removed before determining the wet mass to confirm strogen insufficiency associated with a complete ovariectomy. The mass of the uterus decreases by 7595 in response to ovariectomy. The uteri are then placed in 1095 neutral buffered formalin for subsequent histological analysis.

The right femur is amputated and scanned at the distal metaphysis 1 mm from fissures! of the patella by single-photon absorptiometry. The results of densitometric measurements represent the calculation of bone density depending on the mineral content and width! bones

Ovariectomy rats show a decrease in femur density by 2595 compared to the intact control group treated with the vehicle. Estrogen administered in the orally active form of ethinyl estradiol (392) prevents this bone loss in a dose-dependent manner, but it also has a stimulating effect on the uterus, resulting in uterine mass approaching that of intact rats at dose of 100 μg/kg.

Histological parameters of the uterus.

An increase in height! Zpitelia is a sign of strogennosti therapeutic agents and may be connected with the increased spread of uterine cancer. It is customary to compare the increase in height! compared to the ovariectomized control group. Treatment with estradiol increases the amount of swelling to a thickness greater than that of intact mice. Strogennost is also determined by assessing the adverse reaction of the penetration of the zosinophil into the stroma layer! uterus Estradiol, as expected, causes a large increase in zosinophil penetration.

They also measured the thickness of the streams! and myometrium. Estrogen shows an increase in both of these parameters. A complete definition of strogenity is possible by the sum of all four parameters.

Some data can be described as the percentage of inhibition of bone loss and the percentage of increase in uterine mass, which are calculated as follows:

The percentage of inhibition of bone loss is equal to (bone density of treated Ovex animals - bone density of untreated Ovex animals) and (bone density of estrogen-treated Ovex animals - bone density of untreated Ovex animals) x 100. percentage of mass gain! of uteri is equal to (the mass of the uterus of treated Ovex animals - the mass of the uterus of Ovex animals) and (the mass of the uterus of estrogen-treated Ovex animals - the mass of the uterus of Ovex animals) x 100.

The level of lipids in the serum.

Seventy-five-day-old female Sprague Dawley rats (weight range: 200-225 g) were obtained from Charles River Lzborztoris (Portage, MI). The animals were subjected to either bilateral ovariectomy (0UH) or the Shzma surgical procedure at Charles Lzborztoris and transported one week later. After arrival, the animals were kept in hanging metal cages in groups of 3 or 4 per cage and given as much food as they wanted (calcium content approximately 0.595) and voila! within one week. The temperature in the room was maintained at 22.24-1.72C with a minimum relative humidity of 4095. The light regime in the room was 12 hours of light and 12 hours of darkness!.

Dosing mode/Tissue collection.

After a one-week period of acclimatization (two weeks after OUH), daily dosing of the test compound was started. All compounds were administered orally in the amount of 1 ml/kg of body weight! bodies 17 p-Estradiol was administered subcutaneously in 2095 polyethylene glycol solution as a carrier, 17 c - ethynyl estradiol, and the tested compound was given orally as a suspension in 195 carboxymethyl cellulose or 2095 cyclodextrin. Animals were dosed daily for 4 days. Following the dosing regimen, the animals were weighed and anesthetized with a mixture of Ketamine: Xylazine (2:1 v/v) and a blood sample was collected by cardiac puncture. The animals were then killed by carbon dioxide asphyxiation, the uterus was removed through an incision and the wet mass was determined.

Cholesterol analysis.

Blood samples were allowed to clot at room temperature for 2 hours, and serum was obtained by subsequent centrifugation for 10 minutes at 3000 rpm. Serum cholesterol was determined using a high-characteristic cholesterol assay by Behringer Mannheim Diagnostics. In short, cholesterol was oxidized to cholest-4-en-3-one and hydrogen peroxide. Hydrogen peroxide was then introduced into the reaction with phenol and 4-aminophenazone in the presence of peroxidase! to obtain p-quinonimine dye, which is easy to determine spectrophotometrically at 500 nm. Cholesterol concentration was then calculated according to the standard curve. The entire analysis is automated using the Viomek device

Automated Workshop.

Analysis of zosinophil peroxidase (ZPO) of the uterus.

Matrices are kept at 42C until enzymatic analysis. The uterus is homogenized in 50 volumes of 50 mM Tris buffer (pH 8.0) containing 0.00595 Triton X-100. After adding 0.0195 hydrogen peroxide and 10 mM o-phenylenediamine (final concentration) to the Tris buffer for one minute! an increase in absorption at 450 nm is observed. The presence of zosinophils in the uterus is an indication of estrogenic activity of the compound. The maximum speed in a 15-second interval was set on the initial, linear section of the reaction curve.

Experimental groups.

All experimental groups contained 5-6 animals. Ovariectomy in rats resulted in an increase in serum cholesterol compared to intact vehicle-treated control animals. Estrogen administered in the orally active form of sterinyl estradiol (392) shows a dose-dependent decrease in serum cholesterol, but it also exerts a stimulating effect on the uterus, resulting in uterine mass approaching that of intact rats at a dose of 100 μg /kg/day.

Histological parameter!.

Histological evaluations are performed as described above.

The inhibitory activity of compounds on estrogen-dependent tumors of mammals is evaluated according to the methods described in US patents MoMo 4133814 and 4418068.

The compounds used in the methods of the present invention are effective! in wide dose intervals. For example, the daily dose! will be in the range of 10-1000 mg/kg of mass! bodies For the treatment of adults, the preferred interval is 50-600 mg/kg in the form of a single or divided dose. However, it should be understood that the amount of compounds actually administered will be determined by the doctor depending on the circumstances, including the condition to be treated, the choice of the compound to be administered, the age, weight and response of the individual patient, the severity of the patient's symptoms, and the chosen route of administration. Thus, the above interval! doses do not limit the scope of the invention. While the present compounds are preferably administered orally, they may also be administered by various other routes, such as transdermal, subcutaneous, intranasal, intramuscular, and intravenous routes.

Although it is possible to use the compounds of the invention directly, it is preferable to use them in the form of pharmaceutical preparations containing a pharmaceutically acceptable carrier, diluent or filler and the compound of the invention. Such a drug! will contain from 0.01 to 9995 compounds of the invention.

When preparing the preparations of the present invention, the active component will be casually mixed with at least one carrier, or diluted with at least one carrier, or placed inside the carrier in the form of a capsule, pouch, pocket or second container. If the carrier serves as a diluent, it can be a solid, semi-solid or liquid material acting as a carrier, filler or medium for the active component. So prepare! they can be in the form of tablets, granules, pills, powders, cakes, bags, starch capsules, elixirs, emulsions, solutions, syrups, suspensions, aerosols (solid or in a liquid medium) and soft and hard gelatin capsules.

Try it! suitable carriers, diluents and excipients include lactose, dextrose, sucrose, sorbitol, mannitol, starch, gum arabic, calcium phosphate, alginates, liquid paraffin, calcium silicate, microcrystalline cellulose, polyvinylpyrrolidone, cellulose, tragacanth, gelatin, molasses, me - methylcellulose, methyl- and propylhydroxybenzoate, vegetable oils, such as olive oil, organic substances suitable for injections, such as methyl oleate, talc, magnesium stearate, water and mineral oils. The drug! they can also include wetting agents, lubricants, emulsifiers and suspending agents, preservatives, sweeteners, fragrances, stabilizers, and morsels. Prepare! can be formulated in such a way as to ensure rapid, prolonged or delayed release of the active component after administration to the patient using well-known methods.

For oral administration, the compound of the invention can be perfectly mixed with carriers and diluents and formed into tablets or enclosed in a gelatin capsule!

The drug! preferably made in the form of a standard dose, and each dose contains 1-500 mg, generally 5-300 mg, of the active component. The term "standard dosage form" refers to physical discrete units suitable as single doses for humans and other mammals, and each unit contains a predetermined amount of active material calculated to produce the desired therapeutic effect, mixed with a suitable pharmaceutical carrier, diluted glue or filler.

For a more complete illustration of the invention, here are the following examples! drugs Try it! are only illustrative and do not limit the scope of the invention. The preparation! any of the anti-aging compounds described above can be used as active compounds.

Drug 1.

Hard gelatin capsule! obtained using the following component!:

Quantity Mass concentration per capsule centration (5)

Active component 250 mg 55.0

Dry starch 220 mg 43.0

Magnesium stearate 10 mg 2.0 460 mg 100.0

The above components! mix and place in a hard gelatin capsule in the amount of 460 mg.

Drug 2.

Capsules, each of which contains 20 mg of medication, are made as follows:

Quantity Mass concentration per capsule centration (5)

Active component 20 mg 10.0

Starch 89 mg 44.5

Microcrystalline cellulose 89 mg 44.5

Magnesium stearate 2 MG 1.0 200 mg 100.0

The active component, cellulose, starch and magnesium stearate are mixed, sieved through a US Mo mesh sieve and placed in a hard gelatin capsule.

Drug 3.

A capsule, each of which contains 100 mg of the active component, is obtained as follows:

Quantity Mass concentration per capsule centration (5)

Active component 100 mg 29.0

Polyoxyethylene sorbitan monooleate 50 mg 0.02

Starch powder 250 mg 71.0 250.05 mg 100.02

Bringing the components! mix thoroughly and place in an empty gelatin capsule.

Drug 4.

Tablets, each of which contains 10 mg of the active component, are obtained as follows:

Quantity Mass per capsule concentration (5)

Active component 10 mg 10.0

Starch 45 mg 45.0

Microcrystalline cellulose 35 mg 35.0

Polyvinylpyrrolidone (in the form of 1095 solution in water) 4 mg 4.0

Sodium carboxyethyl starch 4.5 mg 4.5

Magnesium stearate 0.5 mg 0.5

Talc 1 mg 1.0 100 mg 100.0

The active component, starch and cellulose are sieved through a Mo 45 mesh USA sieve and thoroughly mixed. A solution of polyvinylpyrrolidone is mixed with the resulting powder and the mixture is sieved through a Mo 14 mesh USA sieve. Forming granules! dried at 50-602C and sifted through a sieve

Mo 18 mesh USA. Then sodium carboxymethyl starch, magnesium stearate and talc are added to the granules, pre-sifted through a Me 60 mesh USA sieve, mixed and pressed on a tablet machine to obtain tablets weighing 100 mg.

Drug 5.

Tableted drug! can be obtained using the following components!:

Amount - Mass per concentration - capsule ratio (5)

Active component 250 mg 38.0

Microcrystalline cellulose 400 mg 60.0

Silica powder 10 mg 1.5

Stearic acid 5 MG 0.5 665 mg 100.00

Component! mix and press in the form of tablets, each weighing 665 mg.

Drug 6.

Suspensions, each of which contains 5 mg of medication in a dose of 40 ml, are obtained as follows:

For 5 ml of suspension

The active component is 5 mg

Sodium carboxymethyl cellulose 50 mg

Molasses 1.25 ml

Benzoic acid solution 0.10 ml

See how much the flavoring agent absorbs

The dye will see how much it absorbs

How much water is needed up to 5 ml

The drug is sifted through a 45-mesh USA sieve and mixed with sodium carboxymethyl cellulose and molasses to form a homogeneous paste. A solution of benzoic acid, a flavoring agent, and a dye are diluted with a certain amount of water. and add when mixed. Then add a sufficient amount of water! to achieve the required volume.

Drug 7.

An azrosol solution containing the following components is prepared:

Mass concentration (b)

The active component is 0.25

Ztanol 29.75

Propellant 22 (chlorodifluoromethane) 70.00 100.00

The active compound is mixed with ethanol and the mixture is added to the part of the propellant 22, cooled to -302C, and transferred to the filling device. The required amount is then loaded into a stainless steel container and further diluted with the remaining amount of propellant. Then the container is equipped with a valve.

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